Cadherins play critical roles during embryonic development. Extracellularly, they mediate specific cell-cell adhesion, while intracellularly they interact with the catenins and through them with actin cytoskeleton. There is growing evidence that signal transduction via cadherins and catenins induces a variety of cellular responses. In cardiac muscle, N-cadherin has been shown to be important for maintaining structure and function, including myofibril organization and myocyte contraction. The goal of this project is to establish a role for N- cadherin in early events of cardiac muscle formation. Our initial results indicate that N-cadherin is required for maintaining the adhesion between cardiac myocytes of the early mouse embryo. We propose that N-cadherin- mediated cell adhesion regulates intercellular communication and myofibril formation in the early myocardium. We will analyze the ability of cardiac myocytes from N-cadherin-deficient transgenic mouse embryos to assemble gap junctions and desmosomes. The expression and cellular localization of contractile proteins will also be examined in N-cadherin-minus cardiac myocytes. N-cadherin double knockout (DKO) embryonic stem (ES) cell lines will be established. After transfection with GATA-4, the N-cadherin-null ES cells will be differentiated into cardiac muscle in vitro. The ability of epithelial (E-) cadherin to rescue the N-cadherin-deficient cardiomyocytes will be determined. The DKO ES cell line will be transfected with mutant N-cadherins to determine which domains are required for its proper function in cardiac myocytes. Mutant N-cadherins include functionally altered, chimeric, and truncated molecules. Lastly, we will determine if upregulation of N-cadherin is compatible with normal cardiac development. The proposed research will elucidate N-cadherin's role in myocardium formation. In broader terms our results will contribute to our understanding of how cadherins regulate cell morphology and behavior. They also will shed light on the specificity of cadherin function in a particular tissue, in this case cardiac muscle.